High concentrations of calcium suppress osteogenic differentiation of human periodontal ligament stem cells in vitro

BACKGROUND/PURPOSE: Periodontal ligament stem cells (PDLSCs)-based regeneration therapy has received attention for its potential alternative applications in hard tissue and tooth. However, the environmental diversity of oral cavity that regulates PDLSCs differentiation has made it difficult to develop. Therefore, we investigated how high calcium concentrations in the oral environment influence osteogenic differentiation of human PDLSCs (hPDLSCs). MATERIALS AND METHODS: hPDLSCs collected from human molars were isolated and cultured with CaCl(2). First, multi lineage differentiation potentials to osteogenic, chondrogenic, and adipogenic cells were investigated. Then, the effects of CaCl(2) on both alkaline phosphatase (ALP) activity and bone mineralization were analyzed and the expression of mRNA and protein for osteogenic marker was explored. Further, luciferase assay was performed to evaluate CaCl(2) could regulate the transcriptional activity on osteogenic differentiation in hPDLSCs RESULTS: CaCl(2) treatment at normal to high concentrations showed similar suppression of ALP activity, while mineralized nodule formation was decreased by CaCl(2) treatment dose-dependently without affecting proliferation or cytotoxicity in hPDLSCs. We also observed that CaCl(2) treatment repressed the mRNA expression and protein abundance of osteogenic genes and transcriptional factors. Notably, repression of the Runx2 level was significant, and CaCl(2) treatment inhibited Runx2-mediated transcriptional activity on the osteoblast-specific element (OSE) and ALP promoters. CONCLUSION: High concentrations of calcium negatively regulate osteogenic differentiation of hPDLSCs, by repressing osteogenic gene expressions and transcriptional activity. Therefore, these conditions may be applicable to determine the physiologically appropriate concentration of calcium.